P321: PERTURBATIONS IN STIMULUS-INDUCED CYTOKINE RESPONSES IN CHILDREN WITH BCP-ALL
Authors: N. Ruechel, M. Oldenburg, S. Janssen, A. Pandyra, L. Wei, D. Hein, V. Jepsen, U. Fischer, J. Hauer, F. Auer, A. Beer, O. Adams, C. Mackenzie, M. Jaeger, M. Netea, A. Borkhardt, K. Goessling
Journal: HemaSphere (2022)

Background: B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most common subtype of childhood leukemia with a peak during early childhood when children are exposed to viral infections. About 5% of all newborns carry a preleukemic clone, but only about 0,2% of them develop BCP-ALL later in life. It has been assumed that a dysregulated, altered immune response underlies the outgrowth of a pre-leukemic clone in infection-triggered B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Aims: We investigated whether children with BCP-ALL differ in their cytokine response from healthy, age-matched, non-leukemic children, after immune cells were challenged with viral, fungal or bacterial stimuli. Methods: We set up a functional experimental platform with 73 stimulus-cytokine pairs to comprehensively characterize the immune profile of pediatric patients with BCP-ALL with the two most common genetic subtypes, either carrying the ETV6/RUNX1 (E/R) gene fusion or the high-hyperdiploid (HD) karyotype in comparison with an age-and gender-matched healthy control cohort without BCP-ALL and their healthy parents (in total n = 101 individuals). Children were in stable first remission, at least two years after the end of therapy and had a fully reconstituted immune system. Blood was taken of patients and controls in parallel and peripheral blood mononuclear cells (PBMCs) were isolated using a density gradient centrifugation. Cells had been stimulated with various fungal, bacterial and viral pathogens, such as Candida albicans, Staphylococcus aureus, Influenza virus, Respiratory Syncytial virus and toll-like receptor ligands (TLR), such as lipopolysaccharide (TLR4), R848 (TLR7/8) or CpG (TLR9); subsequently cytokines (IFNa, IL-1b, IL-1Ra, IL-10, IL-12p70, IL-17, IFNg) had been measured in the cell culture supernatant. Results: Cytokine base line level did not differ between the groups, while the challenge with various infectious antigens resulted in an overall elevated cytokine production of the BCP-ALL patients with E/R fusion (n=11) (p < 0.01, Mann-Whitney-Wilcoxon test two-sided with Benjamini-Hochberg correction). By deciphering the characteristics of the immune response, a remarkable difference for the anti-viral immune response (p < 0.01) was identified, while the anti-bacterial and anti-fungal response did not differ: The IFNa-response induced by Influenza virus and R848 and the IL-17-response induced by R848 differed most prominently, although not significantly after correction for multiple testing. Further analysis of the cytokine profile of the E/R parents compared with an age- and gender-matched healthy control group revealed an overall significantly elevated cytokine response (p < 0.05) after stimulation, but the different sub-features, such as the fungal, bacterial and viral immune responses were not different. Most strikingly, we could identify this pattern specifically for the E/R cohort. Comparing the cytokine responses of the HD patients (n=8) with the healthy controls (n=22) did not result in any significant differences. Image:Summary/Conclusion: Our study suggests that an altered pro-inflammatory anti-viral cytokine response pattern in children with E/R-BCP-ALL is specific to those children, who later developed the BCP-ALL. It is still present years after chemotherapy and possibly contributes to the outgrowth of a pre-leukemic clone, consistently generated in utero during fetal hematopoiesis.